Tetracyclines (TCs) have an anticollagenase effect; an action unrelated to their antimicrobial property. Recently, it was shown that TCs also inhibited extracellular osteoblastic collagenase (C'ase) activity. However, apart from this anticollagenase action, TCs have another novel property as potent inhibitors of osteoclast function. Therefore, TCs and a new generation of chemically modified nonantimicrobial analogs (CMTs), have enormous therapeutic potential for the treatment of the human lytic bone diseases. This application has a major focus: to define the mechanisms of action for the therapeutic uses of TCs where bone resorption is clinically significant. The specific aims include the following in vitro studies with TCs/CMTs: (1) to examine the transcription, synthesis (intracellular pathways), and activity (extracellular) of the matrix metalloproteinases (MMPs), C'ase and gelatinase (G'ase), with osteoblasts (OBs) and osteoclasts (OCs); (2) to examine the transcription, synthesis, and activity of the TIMPs (tissue inhibitors of the MMPs) with OBs and OCs; (3) to characterize OC reactive oxygen species (ROS); and to investigate the potential of TCs/CMTs to block the extracellular activation of proMMPs from OBs and OCs with or without ROS; (4) to assess the potential inhibitory potency of the new generation CMTs on parameters of OC function; and (5) to determine whether TCs interact with the OC calcium "receptor". cDNA probes to C'ase, G'ase, TIMPs 1 and 2, and the lysosomal enzymes (cathepsin L and TRAP) will measure steady state mRNA levels (transcription). Antibodies will measure synthesis of MMPs and TIMPs. Functional assays will assess the inhibitory action of TCs/CMTs on extracellular MMPs, TIMPs, and lysosomal enzymes. The resorption pit assay will determine the extent of bone resorption with or without TCs/CMTs. ROS will be assayed in OCs with established techniques. To determine whether TCs interact with the OC calcium "receptor", studies will include immunocytochemistry, cytosolic calcium measurements, autoradiography, and morphometry. The proposed studies should provide additional insights into the mechanisms underlying the lytic bone diseases, including periodontitis.